Single-cell sequencing can be expensive, especially compared to conventional bulk RNA sequencing methods. But there is a good reason why this technology is expensive, and you can mitigate these costs if you carefully consider the design of your experiment.
This post looks at the factors affecting the cost of your single-cell sequencing project, and how adapting the design of your experiment can keep costs down.
The costs of the reagents
For a single-cell sequencing experiment, we process single cells in individual reactions, each of which requires its own set of reagents and this drastically increases the total amount of reagents needed.
In contrast, when we analyze pooled cells in a single tube for a bulk RNA-seq experiment, we use much less reagent per cell. Generally speaking, the reagent costs are 10-20 times higher for a standard single-cell RNA sequencing experiment than that of a bulk RNA experiment.
For single-cell sequencing, reagents costs are 10 to 20 times higher than for traditional bulk RNA sequencing.
The cost of sequencing
The cost of sequencing itself is often overlooked when considering the price of a single-cell sequencing project.
Just as with the reagents needed for library preparation, you will need many times more sequencing reads for a standard single-cell sequencing project than for a standard bulk RNA sequencing project.
A typical bulk RNA sequencing analysis requires up to 20 million sequencing reads per sample. In contrast, the typical number of reads per cell needed for a single-cell sequencing project ranges from 50,000 to 150,000*.
So, for example, a relatively simple single-cell sequencing project on the 10x Genomics platform, where you target 3,000 cells with a sequencing depth of 50,000 sequencing reads per cell, will need to generate 3,000 x 50,000 = 150 million reads. That's 7.5 to 15 times more than for a bulk RNA analysis!
Sequencing costs for single-cell sequencing are approximately 7 to 15 times higher than for traditional bulk analyses.
But could you use fewer reads per cell?
Sure, you could. But to gain a thorough understanding of the transcriptome at the level of the individual cell, you need adequate sequencing depth to get a statistically significant dataset.
This is one of the most important trade-offs to consider if you want to keep the sequencing costs stable. If you double the number of cells in your analysis, you will have to halve the number of reads per cell.
The biological question you want to answer will determine which factor is most important.
High-throughput single-cell sequencing is performed on the 10x Chromium X
The technology
Single-cell sequencing is high-tech. It’s a relatively new technology that only recently became available to a larger market thanks to a reduction in costs, as well as innovations in scaling up and standardization.
But, whatever platform or technology you choose, it is more sophisticated than conventional molecular biology technologies such as bulk RNA sequencing.
In order to be able to offer single-cell sequencing in a commercially viable way, we needed to scale down the volume of the reagents used. This could only be done with sophisticated and expensive liquid handling robots.
The same goes for the microfluidics devices that use beads. These beads are microscopic and each of them needs to be coated with a molecular barcode. For one sample, up to 3 million unique barcodes are required. Furthermore, these beads have to run through a microfluidics device smoothly and consistently.
All this requires active research and development, which all adds to the cost of single-cell sequencing.
The setup of your experiment
The most important factor in the cost of single-cell experiments is your biological question. The ultimate price of a project depends on 3 important factors: the number of samples, the number of target cells per sample, and the number of reads per cell.
